Beta-amyloid deposits and neurofibrillary tangles (NFTs) are signature lesions of Alzheimer's disease (AD), but the significance of these lesions is enigmatic. The deposition of Beta-amyloid or A4 peptides (BetaA4) in the AD brain may reflect deranged processing of 1 or more of 3 major CNS amyloid precursor proteins (APPs), and neurons ar a potential source of BetaA4 in AD. To understand the role of human CNS neurons in BetaA4 deposition, it is critical to develop in vitro and in vivo models of normal and aberrant APP metabolism in CNS neurons. New procedures to obtain >99% pure human neurons (NT2N cells) that secrete abundant BetaA4 following treatment of a human teratocarcinoma cell line (Ntera2 or NT2 cells) with retinoic acid now allow studies of APP metabolism and the generation of BetaA4 in human CNS neurons. Since grafts of NT2N cells survive in rodent brains for >1 year, APP biology can be studied in human neurons in an in vivo CNS environment. Finally, injections into rat brains of abnormal tau (i.e. A68) from paired helical filaments (PHFs) in AD NFTs result in co- deposits of BetaA4/A68 interactions in vivo. To accomplish the goals of this Program Project, an Administrative and a Neuroscience Core facilitate the research of 4 Projects. Project 1 studies the biosynthesis, processing, intracellular transport and recycling of APPs as well as the site(s) at which BetaA4 is generated in neurons and secreted using wild type and transfected NT2/NT2N cells. Project 2 characterizes APP glycoforms from AD and control brains and determines the role of glycosylation in APP metabolism and in the production of BetaA4 in NT2/NT2N cells and in glycosylation deficient non-neuronal cells transfected with APP CDNAS. Project 3 examines interactions of APPs with the neuronal cytoskeleton in NT2/NT2N cells and assesses effects of perturbations of the neuronal cytoskeleton on APP metabolism and BetaA4 production. Finally, Project 4 models BetaA4 pathology by grafting NT2N cells, or by injecting NFT components (e.g. A68 derived from AD brains) into the CNS of rodents. In sum, this Program Project will develop a more precise understanding of APP metabolism and mechanisms leading to the production of BetaA4 in human neurons, delineate critical metabolic events that lead to the deposition of BetaA4 in the AD brain and establish in vitro and in vivo models of AD amyloidosis.